Increased mechanical stress in the heart under pathological conditions such as hypertension, infarction and fibrosis can cause arrhythmias and heart failure. However, little is known about the mechano-chemo transduction mechanisms that underlie heart disease development due to a previous lack of practical techniques to control mechanical loading on single, intact cardiomyocytes necessary for investigating at cellular and molecular levels. Recently, we developed a novel Cell-in-Gel system that allows for the control of mechanical loads on single cardiomyocytes (freshly isolated from healthy adult rabbit hearts and from heart failure disease model) in a 3D viscoelastic gel matrix. We will use this system to study the mechanical loading effects on modulating myocyte calcium signaling and contraction dynamics during excitation-contraction coupling. The goal of this project is to use the Cell-in-Gel system and mechanical model analysis methods as tools to study mechano-chemo transduction mechanisms in intact rabbit cardiomyocytes. I plan to engage in an interdisciplinary venture to achieve 2 specific aims. Aim 1: I will implement the Cell-in-Gel system to investigate biomechanics involved in mechano-chemo transduction at the single-cell level in cardiomyocytes from healthy control rabbits. I will specifically look at the effects of preload and afterload on the contraction and calcium signaling systems. Calcium signaling will be measured using epifluorescence and confocal microscopy methods to quantify systolic and diastolic calcium. Contraction will be measured concurrently with calcium using video-based methods. We predict increased mechanical loading will cause compensatory augmentation of calcium signaling to decreased contraction, contributing to arrhythmias and heart disease. Aim 2: I will perform similar experiments in Aim 1 on cells from a heart failure rabbit model. We expect mechano-chemo transduction to be attenuated in heart failure cells due to calcium dysregulation originating from pathophysiological changes in rabbit hearts experiencing chronic, excessive mechanical loading in vivo. Understanding mechano-chemo transduction mechanisms at the cellular level can provide the basis for translation to the tissue and organ level. Identifying the specific mechano-chemo transduction pathways involved in cardiac pathologies will reveal opportunities for therapeutic approaches for treating hypertension, arrhythmia and heart failure.